The overall objectives of this project are to gain insights into the role of the growth factor receptor, Mpl, in hematopoiesis and to provide in-depth training in Molecular Biology for Dr. Diana Sabath. The purpose of this training is to enable Dr. Sabath to achieve her immediate career goal of studying Mpl biology and her long-term career goal of becoming an independent Clinical Scientist involved in laboratory-based research at a major academic medical center. To achieve these goals, Dr. Sabath will follow a research career development plan consisting of a program of didactic sessions and a laboratory-based research project under the co-sponsorship of Dr. Virginia C. Broudy and Dr. Kenneth Kaushansky. The objectives of this research project are to understand the structure/function relationships of Mpl and to determine how these relationships contribute to normal and abnormal hematopoiesis. The specific aims of this research project are to: 1) identify components of the Tpo/Mpl receptor complex; 2) define the role of the duplicated extracellular domains of murine Mpl; and 3) determine if different forms of Mp1 serve different functional roles. The research design and methods for this project include characterization of the Tpo/Mpl receptor complex using receptor binding studies and affinity crosslinking, functional analysis of the extracellular domain of Mpl by creating altered forms of Mpl using PCR-mediated mutagenesis and DNA cloning for subsequent stable transfection into growth factor responsive cell lines, and comparison of the sequences of c-mpl cDNA obtained from three different cell lines with different physiologic responses to Tpo. The health relatedness of this project is twofold: 1) an abnormal form of Mpl causes myeloproliferative disease in mice; and 2) high levels of expression of Mpl are associated with a poor prognosis in leukemia and myelodysplasia. Study of the structure/function relationships of Mpl may lead to insights into the development of these and other hematologic diseases.